Todesco Hayley M, Gafuik Chris, John Cini M, Roberts Erin L, Borys Breanna S, Pawluk Alexis, Kallos Michael S, Potts Kyle G, Mahoney Douglas J
Arnie Charbonneau Cancer Institute, Faculty of Medicine, University of Calgary, Calgary, AB, Canada.
Alberta Children's Hospital Research Institute, Faculty of Medicine, University of Calgary, Calgary, AB, Canada.
Mol Ther Methods Clin Dev. 2024 Jan 17;32(1):101189. doi: 10.1016/j.omtm.2024.101189. eCollection 2024 Mar 14.
The severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) pandemic highlighted the importance of vaccine innovation in public health. Hundreds of vaccines built on numerous technology platforms have been rapidly developed against SARS-CoV-2 since 2020. Like all vaccine platforms, an important bottleneck to viral-vectored vaccine development is manufacturing. Here, we describe a scalable manufacturing protocol for replication-competent SARS-CoV-2 Spike-pseudotyped vesicular stomatitis virus (S-VSV)-vectored vaccines using Vero cells grown on microcarriers in a stirred-tank bioreactor. Using Cytodex 1 microcarriers over 6 days of fed-batch culture, Vero cells grew to a density of 3.95 ± 0.42 ×10 cells/mL in 1-L stirred-tank bioreactors. Ancestral strain S-VSV reached a peak titer of 2.05 ± 0.58 ×10 plaque-forming units (PFUs)/mL at 3 days postinfection. When compared to growth in plate-based cultures, this was a 29-fold increase in virus production, meaning a 1-L bioreactor produces the same amount of virus as 1,284 plates of 15 cm. In addition, the omicron BA.1 S-VSV reached a peak titer of 5.58 ± 0.35 × 10 PFU/mL. Quality control testing showed plate- and bioreactor-produced S-VSV had similar particle-to-PFU ratios and elicited comparable levels of neutralizing antibodies in immunized hamsters. This method should enhance preclinical and clinical development of pseudotyped VSV-vectored vaccines in future pandemics.
严重急性呼吸综合征冠状病毒2（SARS-CoV-2）大流行凸显了疫苗创新在公共卫生中的重要性。自2020年以来，基于众多技术平台已迅速开发出数百种针对SARS-CoV-2的疫苗。与所有疫苗平台一样，病毒载体疫苗开发的一个重要瓶颈是生产。在此，我们描述了一种可扩展的生产方案，用于生产具有复制能力的SARS-CoV-2刺突蛋白假型水疱性口炎病毒（S-VSV）载体疫苗，该疫苗使用在搅拌罐生物反应器中的微载体上生长的Vero细胞。在6天的分批补料培养中使用Cytodex 1微载体，Vero细胞在1升搅拌罐生物反应器中生长至密度为3.95±0.42×10个细胞/毫升。感染后3天，原始毒株S-VSV达到峰值滴度2.05±0.58×10空斑形成单位（PFU）/毫升。与基于平板培养的生长情况相比，病毒产量增加了29倍，这意味着1升生物反应器产生的病毒量与1284个15厘米平板产生的病毒量相同。此外，奥密克戎BA.1 S-VSV达到峰值滴度5.58±0.35×10 PFU/毫升。质量控制测试表明，平板和生物反应器生产的S-VSV具有相似的颗粒与PFU比率，并在免疫仓鼠中引发了相当水平的中和抗体。该方法应能加强未来大流行中假型VSV载体疫苗的临床前和临床开发。
